CGI Italia: DNA-FISH Probe Catalog

DNA-FISH Probe CAtAlog

NHl

AMlAlCl All Cll

CMl MM MDS

Hematologic Diseases

Solid tumors

breast

lung

Cervical

Colorectal

[email protected] 2013

EXCELLENCE IN ONCOLOGY

Visit our website, www.cancergeneticsitalia.com, and keep abreast of our latest product releases, access complete information about our product portfolio, and learn about our R&D pipeline.


table of Contents

4 About CgI Italia

5 our Commitment to Quality

6 What is FISH?

8 DNA-FISH Probe Portfolio

28 Appendix

28 Scoring guidance and Signal Interpretation

29Filter Specifications

troubleshooting

33 ordering Products

33 ordering Information

34 Product Index by Disease

36 Product Index by Name

37 Product Index by Chromosome Number

Page 4

about CGi italiaCancer Genetics Italia S.r.l. (CGI Ita-

lia) was founded in 2009 and is headquar-tered in Milan, Italy. CGI Italia is a wholly owned subsidiary of Cancer Genetics Inc. (CGI), which was founded in 1999 by world-renowned human geneticist R.S.K. Chaganti, Ph.D. CGI’s affiliation with major cancer centers along with the firm’s foun-dation in world-class scientific knowledge has enabled CGI to develop strong intellec-tual properties (IP) in solid and blood-borne cancers.

It is from this foundation that CGI Italia draws its ability to design and manufacture proprietary DNA-FISH Probes. Our DNA-FISH Probes detect a range of genetic ab-errations, such as translocations and copy number changes, in several cancer types and are protected by the following patents:

§ U. S. Patent (#7,585,964, Publication: pending) & CAN Patent (2,447,320, Published: 21 November 2002) entitled Methods of Analyzing Chromosomal Translocations using Fluorescence in situ Hybridization (FISH).

§ U. S. Patent (#11/932,422, Publica-tion: 30 April 2009) & EU PCT Patent (#08844570.5, Published: 07 May 2009) entitled Panel for the Detection and Dif-ferentiation of Renal Cortical Neoplasms.

§ U. S. Patent (#13/227,027, Publication: pending & PCT/US2011/050681, Pub-lished: 15 March 2012) entitled Methods for Detecting Human Papilloma Virus-As-sociated Cancers.

Our products are marketed globally and are provided in a ready-to-use format as CE marked or Research Use Only (RUO).

Mission stateMentOur mission is to provide professionals

with robustly designed DNA-FISH Probes that are highly specific, easy to interpret, and possess strong signal quality, for the clinical management of cancer patients.

Milestones 2009• CGI Italia is founded

2010• 12 CE products are launched

2012• Portfolio expands to 31 CE products• Release of FHACTTM (FISH based

HPV-Associated Cancer Test)2013• Release of D13S319

littlefisH ProGraMThrough its LittleFISH Program, CGI Ital-

ia sponsors academic efforts involving FISH technology. This program provides under-graduate, graduate, and training physician students with discounted FISH Probes, and aims to encompass research projects as well as training and certification programs teaching the FISH technique. Laboratory reagents can represent an expense that may be difficult for academic laboratories and training programs to manage. By giv-ing students a “Little” help, CGI Italia hopes to support research and promote FISH technology in the management of cancer around the globe.

For further information, contact CGI Italia at [email protected].

Academic Sponsorship Program

Cancer Genetics Italia2013 DNA-FISH Probe Catalogwww.cancergeneticsitalia.com

Page 5

a new standard for fisH Probes:our CoMMitMent to Quality

“The results of our comparison study demonstrate that CGI Italia Probes are user-friendly, robust and produce signals that are easy to score. The signals are very specific, bright and balanced in intensity, and have low levels of back-ground noise. CGI Italia is a great choice for DNA-FISH Probes.”

Sankar Mohan, Ph.D., OncoMDx Laboratories

CGI Italia takes great care in developing FISH Probes, and ensuring that every lot of Probe meets a quality standard.

• Optimal concentration of each probe is rigourously determined to ensure balance of sig-nal intensity among the 2-4 color probes, as well as minimized background noise.

• Enumeration Probes are paired with a control locus to give the end-user a more complete picture of the gain/deletion of interest. Accurate screening of Enumeration Probes is gen-erally not complete without control loci. Signal patterns of the locus of interest and control locus make it possible to gauge the likelihood of a whole chromosome deletion versus a localized deletion, which, in many cases, is important to distinguish.

• During the validation and manufacture stages, the FISH probes are tested on normal peripheral blood, intended tissue (bone marrow, formalin-fixed paraffin embedded tissue, or liquid cervical specimen) to ensure sensitivity, specificity, and reproducibility of results. Break Apart & Fusion Probes are also tested on positive cell lines to ensure fusion sig-nals are well-balanced.

• With regards to CE versus RUO classification, RUO Probes are subject to the same QA/QC standard as CE-marked Probes as part of CGI Italia’s commitment to providing high quality products.

• Finally, some customers may request cut-off values for positive versus negative FISH results. It should be noted that CGI Italia does not provide cut-off values as it is recom-mended that each laboratory determine its own cut-offs based on testing of normal speci-mens. This is generally a requirement for most clinical laboratories.

• CGI Italia FISH Probes have been designed with the intention of providing robust prod-ucts that outperform competition. For more information about our comparison study con-ducted by OncoMDx, please contact us at [email protected].

Page 6

wHat is fisH?Fluorescence in situ hybridization (FISH) is a sensitive and accurate technique that enables

the detection of chromosomal aberrations and is complementary to conventional cytogenet-ic analysis. The method entails the hybridization of a single-stranded fluorescently labeled nucleic acid sequence (probe), which is complementary to a target genomic sequence that is present in metaphase chromosomes as well as interphase nuclei. Once the hybridization is complete, the excess FISH probe is washed off from the slide, which allows signal localization and enu-meration.

The primary advantage of the FISH technique is its applicability to non-dividing cells and a variety of speci-men types. The technique allows the detection of a given genomic abnormal-ity such as translocation, gain, or loss. FISH is also a preferred method for diag-nosis, prognosis, treatment response, and minimal re-sidual disease detection in a variety of hematopoietic neoplasms and solid tu-mors.

DNA-FISH Probe

PortFolIo

Page 8

disease leGend

AlCl Anaplastic large Cell lymphoma

All Acute lymphoblastic leukemia

AMl Acute Myeloid leukemia

Cll Chronic lymphocytic leukemia

CMl Chronic Myeloid leukemia

MM Multiple Myeloma

MDS Myelodysplastic Syndrome

NHl Non-Hodgkin’s lymphoma


Page 9

table of Contents

105p15, 9q34, 15q24 • Hyperdiploidy 5, 9, 15 • ref: 16-003 MM

A20/PRDM1/SHgC-79576 • del(6q23),del(6q21)/6p12 • ref: 19-001 All

NHl

11ABL1/BCR • t(9;22) • ref: 10-001 All

CMl

ALK break Apart • t(2p23) • ref: 21-002 AlCl

Solid tumor

12ALK/NPM1 • t(2;5) • ref: 21-001 AlCl

AML1/ETO • t(8;21) • ref: 12-005 AMl

13API2/MALT1 • t(11;18) • ref: 17-001 NHl

ATM/D11S1251 • del(11q22)/11p15 • ref: 14-018 Cll

NHl

14BCL6 break Apart • t(3q27) • ref: 18-010 NHl

CCND1/IGH • t(11;14) • ref: 14-006 MM

NHl

15D7S486/Cen7 • del(7q31)/Cen7 • ref: 11-007 AMl

MDS

D13S25/D13S1009 • del(13q14)/13q34 • ref: 14-009 Cll

MM

NHl

16D20S108/8q11 • del(20q12) & trisomy 8 • ref: 11-001 AMl

MDS

EGFR/Cen7 • EGFR Amplification • ref: 22-007 Solid tumor

17EGR1/5p15 • del(5q31)/5p15 • ref: 11-004 AMl

MDS

ERBB2/Cen17 • ERBB2 Amplification • ref: 22-003 Solid tumor

18FGFR3/IGH • t(4;14) • ref: 16-005 MM

FHACttM • 3q26/5p15/20q13/Cen7 • ref: 25-002 Solid tumor

19IGH/BCL2 • t(14;18) • ref: 18-001 All

NHl

IGH break Apart • t(14q32) • ref: 13-014 All

MM

NHl

20IGH/MAF • t(14;16) • ref: 16-010 MM

IGH/MALT1 • t(14;18) • ref: 17-004 NHl

21MDM2/D12S1837 • 12q15/12p11 • ref: 14-017 Cll

NHl

MLL break Apart • t(11q23) • ref: 11-002 All

AMl

22MYB/SHgC-79576 • 6q23/6p12 • ref: 14-016 All

Cll

MYC break Apart • t(8q24) • ref: 13-008 All

Cll

NHl

23MYC/IGH • t(8;14) • ref: 13-004 All

NHl

MYH11/CBFB • inv(16) • ref: 12-010 AMl

CMl

MDS

24PBX1/E2A • t(1;19) • ref: 13-001 All

PML/RARA • t(15;17) • ref: 12-008 AMl

CMl

25RB1/D13S1009 • del(13q14)/13q34 • ref: 27-014 AMl

Cll

MDS

MM

TP53/RARA • del(17p13)/17q21 • ref: 14-015 AMl

Cll

MDS

Solid tumor

Page 10

5p15, 9q34, 15q24Three Color, Enumeration ProbeRef: 16-003

A20/PRDM1/SHgC-79576Three Color, Enumeration ProbeRef: 19-001

The 5p15, 9q34, 15q24 DNA-FISH Probe is designed to detect changes in copy number of chromosome(s) 5, 9, and 15 by fluorescence in situ hybrid-ization (FISH). Hyperdiploidy is characterized by increased copy numbers of chromosomes such that the modal chromosomal number is 47-57. It is ob-served in 30-50% of multiple myeloma (MM) cases where the most common trisomies involve chromosomes 3, 5, 7, 9, 11, 15, 19, and 21.[1] In a diagnostic setting, gain of at least two of the three chromosomes 5, 9, or 15 in MM is considered an indicator for hyperdiploidy.[2] When assessed by G-banded karyotype and in the absence of other aberrations such as IGH transloca-tions or deletion of chromosome 13, hyperdiploidy in MM is associated with a good prognosis.[3-5]

references1. Liebisch, P., Döhner, H. Eur J Cancer, 2006.

42(11): p. 1520-9.2. Chen, L., et al. Exp Oncol, 2007. 29(2): p. 116-20.3. Chng, W. J., et al. Leukemia, 2006. 20(5): p.

807-13.4. Boyd KD, et al. NCRI Haematology Oncology

Studies Group. Leukemia. 2012 26(2): p 349-55. 5. Kumar S, et al. Blood, 2012. 119(9): p 2100-5.

MM

NHl

The A20 (also called TNFAIP3)/PRDM1/SHGC-79576 DNA-FISH Probe is designed to detect deletion of the A20 gene located on 6q23 and the PRDM1 gene located on 6q21 relative to the control locus SHGC-79576 on 6p12, using fluorescence in situ hybridization (FISH). Deletion of 6q is observed in all types of B-cell malignancies, where two commonly deleted regions map to the A20 and PRDM1 gene region.[1,2] Loss of the A20 gene has been observed in ~20% of non-Hodgkin lymphoma (NHL) cases, ~20% of muco-sa-associated lymphoid tissue (MALT) lymphoma cases[3] and in nonsplenic marginal zone lymphomas (MZLs) cases.[4] The loss of the A20 gene has been observed most frequently in mantle cell lymphoma (MCL) and diffuse large B-cell lymphoma (DLBCL) cases with a frequency rate of 31% and 38%, respectively.[5] In DLBCL, inactivation of A20 is seen more frequently in the activated B-cell (ABC) subtype of DLBCL (50%) versus the germinal center B-cell (GCB) subtype (22%).[3,5,6] DLBCL cases exhibit a variety of complex 6q deletions, which encompasse either A20 or PRDM1 alone or to-gether as part of a larger deletion.[2] Additionally, the deletion of the PRDM1 gene has been observed in 53% of primary central nervous system lympho-mas (PCNSLs).[7,8]

references1. Offit, K., et al. Blood, 1993. 82(7)L p. 1781-7.2. Thelander, E. F., et al. Leuk Lymphoma, 2008. 49(3):

p.477-87.3. Kato, M., et al. Nature, 2009. 459(7247): p.712-6.4. Novak, U., et al. Blood, 2009. 113(20): p.4918-21.5. Compagno, M., et al., Nature, 2009. 459(7247): p.

717-21.6. Lenz, G., et al. PNAS, 2008. 105(36): p.13520-5.7. Schwindt, H., et al. Leukemia, 2009. 23(10):

p.1875-84.8. Pasqualucci, L. et al., J Exp Med 2003 (2): p 311-7.

All

9

5p15

AFMA055ZD9

~554 kb

5

centromere telomere 9q34

D9S64

~468 kb

telomere centromere

15q24

D15S169telomere centromere

15

~269 kb

5’ 3’SHgC-79576

~772 kb

6

telomere telomereA20

~494 kb ~774 kb

5’ 3’PRDM1

5p15

, 9q3

4, 1

5q24

A20

/PR

DM

1/SH

gC

-795

76


Page 11

ABL1/BCRTwo Color, Two Fusion Translocation ProbeRef: 10-001

ALK break ApartTwo Color, Break Apart ProbeRef: 21-002

All CMl

AlCl lung

The ALK Break Apart DNA-FISH Probe is designed to detect the transloca-tion between the ALK gene located at 2p23 and one of at least 14 known translocation partner loci using fluorescence in situ hybridization (FISH).[1]

Translocation of the ALK gene occurs in ~50% of anaplastic large cell lym-phoma (ALCL) cases with a form of t(2;5)(p23;q35), as determined by con-ventional cytogenetics; in such cases, the presence of the t(2;5)(p23;q35) form carries a better overall prognosis for ALCL patients.[2] ALK translo-cation has been observed in vesical inflammatory myofibroblastic tumors (IMT) of the bladder (>66%)[3] and serves as a diagnostic biomarker for dif-ferential diagnosis of IMT from sarcomatous lesions.[3] ALK translocation is observed in ~5% - 16% of non-small cell lung cancer (NSCLC) cases in the form of inv(2)(p21p23), as determined by FISH,[4] and serves as a biomarker for therapy response.[4,5] The presence of the ALK translocation in NSCLC patients is correlated with a marked sensitivity to pemetrexed and crizotinib treatment.[6]

references1. Chiarle, R., et al. Nat Rev Cancer, 2008. 8(1): 11-23.2. Drexler, H. G., et al. Leukemia, 2000. 14(9): p.1533-

59.3. Sukov, W. R., et al. Mod Pathol, 2007. 20(5): p.592-

603.4. Kwak, E.L., et al. N Eng J Med, 2010. 363(18): p.

1693-703.5. Gerber, D. E., et al. Cancer Cell, 2010. 18(6): p.548-

51.6. Camidge, D. R., et al. J Thorac Oncol, 2011.

6(4):p.774-80.

The ABL1/BCR DNA-FISH Probe is designed to detect the translocation between the ABL1 gene on chromosome 9q34 and the BCR gene on chro-mosome 22q11 by fluorescence in situ hybridization (FISH). This reciprocal translocation results in the Philadelphia chromosome (Ph), the der(22), and is the hallmark of chronic myeloid leukemia (CML). Approximately 90-95% of CML and up to 5% of pediatric and 20% of adult acute lymphocytic leukemia (ALL) are Ph positive.[1-3] ABL1/BCR FISH is used in diagnosis, prognosis, and monitoring of t(9;22) in CML and ALL patients.[4] A subset of CML (~10%) and ALL (~5%) cases exhibit large deletions adjacent to the breakpoints on chromosomes der(9) and der(22).[4-5] Such submicroscopic losses carry a poor prognosis[6] and can be detected by the Cancer Genetics Italia DNA-FISH Probe.

references1. Huret, J. L. t(9;22)(q34;q11) in CML, Dec. 1997 www.AtlasGeneticsOncology.org. 2. Huret, J. L. t(9;22)(q34;q11) in ALL, Sep 1997 www.AtlasGeneticsOncology.org. 3. Nashed, A. L., et al. J Mol Diagn, 2003. 5:63-72. 4. Landstrom, A.P., Tefferi, A. Leuk Lymphoma, 2006. 47(3): 397-402.5. Gorusu, M., et al. Cancer Genet Cytogenet, 2007. 173:97-106.6. Huntly, B.J. et al. Blood, 2003. 120(4): 1160-8.

2

3’ 5’2p23

ALK

~556 kb

centromere telomere

~541 kb

9

22

telomerecentromereBCR

5’ 3’22q11

~350 kb ~390 kb

centromere telomereASSABL1

5’ 3’9q34

~285 kb ~390 kb

AB

L1/B

CR

ALK

bre

ak A

part

Page 12

ALK/NPM1Two Color, Two Fusion Translocation ProbeRef: 21-001

AML1/ETOTwo Color, Two Fusion Translocation ProbeRef: 12-005

AlCl

AMl

The ALK/NPM1 DNA-FISH Probe is designed to detect the translocation between the ALK gene located at 2p23 and the NPM1 gene located at 5q35, using fluorescence in situ hybridization (FISH); the translocation between the ALK and NPM1 gene is designated as t(2;5)(p23;q35). By convention-al cytogenetics, the translocation occurs in up to 50% of anaplastic large cell lymphoma (ALCL) cases.[1] As assessed by immunohistochemistry, ex-pression of the fusion protein ALK/NPM1 that is generated by t(2;5), occurs more frequently in childhood ALCL with an occurrence rate of 83% versus adult ALCL which has an occurrence rate of 31%.[1,2] The presence of t(2;5)(p23;q35) carries a better overall prognosis for ALCL patients.[1]

references1. Drexler, H.G., et al. Leukemia, 2000. 14(9):

p.1533-59.2. Weitzman, S., et al. Curr Oncol Rep, 2002. 4(2):

p.107-13.

The AML1/ETO (also named RUNX1/RUNX1T1) DNA-FISH Probe is de-signed to detect the translocation involving the AML1 (RUNX1) gene on chromosome 21q22 and the ETO (RUNX1T1/ MTG8) gene on chromosome 8q22 by fluorescence in situ hybridization (FISH). This translocation gener-ates an AML1-ETO fusion protein and is detected in 12% of de novo AML cases and in up to 46% of cases in the AML subtype M2.[1,2] The detection of the t(8;21) is clinically relevant because it is generally associated with a favorable prognosis and particularly with high-dose cytorabine-based con-solidation chemotherapy.[3]

references1. Nucifora, G., Rowley, J.D. Blood (Review), 1995.

86(1):1-14. 2. Peterson, L. F., Zhang, D.E. Oncogene, 2004.

23(24):4255-62. 3. Dohner, H., et al. Blood, 2010. 15(3): 453-74.

3’ 5’2p23

ALK

~556 kb

2

centromere telomere

~541 kb

5

5’ 3’5q35

NPM1

~502 kb

telomere centromere

~552 kb

8

21

ETO3’ 5’8q22

~635 kb ~472 kb

AML13’ 5’21q22

~570 kb ~630 kb

centromere telomere

centromere telomere

ALK

/NPM

1A

ML1

/ETO


Page 13

API2/MALT1Two Color, Two Fusion Translocation ProbeRef: 17-001

ATM/D11S1251Two Color, Enumeration ProbeRef: 14-018

Cll

NHl

The API2/MALT1 DNA-FISH probe is designed to detect the translocation between the API2 gene located at 11q21 and the MALT1 gene located at 18q21 using fluorescence in situ hybridization (FISH).[1] The translocation between the API2 and MALT1 gene, designated as t(11;18)(q21;q21), can be detected in around 15% of mucosa-associated lymphoid tissue (MALT) lym-phomas, but varies in frequency based on primary tumor site.[2] In pulmonal and gastric MALT, t(11;18) is found more frequently (38-53% and 22-24%, respectively) and in these cases is almost always the only detected chromo-somal abnormality.[2] When observed in gastric MALT lymphoma, t(11;18) is highly associated with a lack of response to antibiotic H. pylori eradication treatment.[3,4]

references1. Dierlamm, J., et al., Blood, 2000. 96(6): 2215-18.2. Heim, S., and Mitelman, F. (Ed) Cancer Cytoge-

netics, 2009 (3rd Edition), Wiley-Blackwell, New Jersey. p. 317-320.

3. Nakamura, T., et al. J Gastroenterol, 2003. 38(10): p.921-9.

4. Toracchio, S., et al. Cancer Sci, 2009. 100(5): p.8881-7.

The ATM/D11S1251 DNA-FISH Probe is designed to detect the deletion of the ATM gene located on 11q22 relative to the control locus D11S1251 lo-cated on 11p15 by fluorescence in situ hybridization (FISH). ATM deletions are frequently seen in several types of hematologic malignancies. The dele-tion of the ATM gene is detected in ~65% of T-cell prolymphocytic leukemia (T-PLL) cases[1], ~50% of mantle cell lymphoma (MCL) cases[1,2], and ~20% of chronic lymphocytic leukemia (CLL) cases.[3] Deletion of 11q in CLL pa-tients is associated with extensive lymphadenopathy, disease progression, and shorter median survival.[3,4] Significantly improved clinical outcomes in previously untreated CLL patients with ATM loss have been observed using alkylating agent-based chemo-immunotherapy regimens.[5]

references1. Monni, O., Knuutila, S. Leuk Lymphoma, 2001.

40(3-4): p. 259-66.2. Stilgenbauer, S., et al., Blood, 1999. 94(9): p. 3262-4.3. Dohner, H., et al. N Engl J Med, 2000. 343(26): p.

1910-6.4. Stilgenbauer, S., et al., Leukemia, 2002. 16((6): p.

993-1007.5. Tsimberidou, A.M., et al. Cancer, 2009. 115(2): p.

373-80.

NHl

11

11q21

API2

~645 kb

telomere centromere

~498 kb

18

5’ 3’18q21

MALT1

~738 kb

telomere centromere

~641 kb

5’ 3’

5’ 3’ATM

~399 kb

D11S1251

~423 kb

11

telomere telomere

API

2/M

ALT

1AT

M/D

11S1

251

Page 14

BCL6 break ApartTwo Color, Break Apart ProbeRef: 18-010

CCND1/IGHTwo Color, Two Fusion Translocation ProbeRef: 14-006

NHlMM

The CCND1/IGH DNA-FISH Probe is designed to detect the translocation between the CCND1 gene located on 11q13 and the IGH gene located on 14q32 by fluorescence in situ hybridization (FISH). The translocation be-tween the CCND1 and IGH gene is designated as t(11;14)(q13;q32) and is the cytogenetic hallmark of mantle cell lymphoma (MCL), which distinguishes it from other non-Hodgkin lymphomas.[1,2] The t(11;14) also has been detected in light chain amyloidosis, monoclonal gammopathy of undetermined sig-nificance (MGUS), and multiple myeloma (MM) cases.[3,4] The presence of t(11;14)(q13;q32) in MM patients is associated with an improved survival.[5]

references1. Heim, S., Mitelman, F. (Ed). Cancer Cytogenetics,

2009 (3rd Edition). Wiley-Blackwell, New Jersey. p. 313-315.

2. Bentz, J.S., et al. Cancer, 2004. 102(2): p.124-31.3. Barille-Nion, S., et al. Hematology Am Soc Hematol

Educ Program, 2003. p.248-78.4. Liebisch, P., et al. Eur J Cancer, 2006. 42(11):

p.1520-9.5. Fonseca, R., et al. Leukemia, 2009. 23(12): p.2210-

21.

NHl

The BCL6 Break Apart DNA-FISH Probe is designed to detect the transloca-tions between the BCL6 gene located on 3q27 and one of at least 20 known translocation partner loci as detected by fluorescence in situ hybridization (FISH). Translocation of the BCL6 gene occurs in 6-26% of follicular lympho-ma (FL)[1] with higher incidence (44%) in grade 3 cases negative for t(14;18)(q32;q21).[2] Rearrangement of the BCL6 gene is observed at a frequency of 15~40% in diffuse large B-cell lymphomas (DLBCL).[1-3]

references1. Chaganti, R.S., et al. Semin Hematol, 2000. 37(4):

p. 396-411.2. Gu, K., et al. Mod Pathol, 2009. 22(9): p. 1251-7.3. Lo Coco, F., et al., Blood, 1994. 183(7): p. 1757-9.

3’ 5’ 3q27

BCL6

~756 kb

3

telomere centromere

~471 kb

14

11q13

CCND1

11

~511 kb~513 kb

telomere centromere 5’ 3’

3’14q32

~561 kb ~420 kb

5’C segments J s

egme

nts

D se

gmen

ts

V segmentsIGH

centromere telomere

BC

L6 b

reak

Apa

rtC

CN

D1/

IGH


Page 15

D7S486/Cen7Two Color, Enumeration ProbeRef: 11-007

D13S25/D13S1009Two Color, Enumeration ProbeRef: 14-009

AMl MDS

Cll

MM

The D7S486/Cen7 DNA-FISH Probe is designed to detect the deletion of the D7S486 locus located on 7q31 relative to the centromere 7 control locus, us-ing fluorescence in situ hybridization (FISH). Deletions of chromosome 7 are frequently seen in many types of myeloid neoplasms. Loss of the D7S486 locus is detected in ~5% of adults with de novo myelodysplastic syndrome (MDS) and in ~50% of children with de novo MDS.[1] Additionally, loss of the D7S486 locus is observed in 5% of de novo acute myeloid leukemia (AML) patients and in 30 - 40% of therapy related MDS (t-MDS)/AML (t-AML) pa-tients.[1,2] Deletion of the D7S486 locus or monosomy of chromosome 7 is associated with a poor prognosis in adults and children diagnosed with MDS and/or AML.[1,3]



2. Shali, W., et al., Cancer Genet Cytogenet, 2006. 168(2): p. 133-145.

3. Haase, D. Ann Hematol, 2008. 87(7): p.515-26.

The D13S25/D13S1009 DNA-FISH Probe is designed to detect loss of the D13S25 locus on chromosome 13q14 relative to the control marker D13S1009 on chromosome 13q34 by fluorescence in situ hybridization (FISH). The de-letion of D13S25, a locus distal to the RB1 gene, has been detected in B-cell chronic lymphocytic leukemia (B-CLL),[1] in multiple myeloma (MM),[2,3] and rarely in a variety of non-Hodgkin lymphomas (NHL).[4] Deletion of 13q14 has a strong prognostic value correlating with slower disease progression and better prognosis in B-CLL patients,[5] whereas in MM patients it is associated with a higher stage of disease and shorter survival.[6]

references1. Nelson, B. P., et al. Am J Clin Pathol, 2007. 128(2):

323-32.2. Chen, L., et al. Exp Oncol, 2007. 29(2):116-20.3. Terpos, E., et al. Leuk Lymphoma, 2006. 47(5):803-

14.4. Dierlamm, J., et al. Cancer Genet Cytogenet, 2000.

1;120(1):1-5.5. Dal Bo, M., et al., Genes Chromosomes Cancer,

2011. 50(8): p. 633-43.6. Kroger, N., et al., Blood, 2004. 103(11): p.4056-61.

NHl

7q31

D7S486

~230 kb

7

telomere centromere

Cen7

centromere telomere

~186 kb ~321 kb

D13S1009

13

D13S25

D7S

486/

Cen

7D

13S2

5/D

13S1

009

Page 16

D20S108/8q11Two Color, Enumeration ProbeRef: 11-001

EGFR/Cen7Two Color, Enumeration ProbeRef: 22-007

AMl MDS

The D20S108/8q11 DNA-FISH Probe is designed to detect the deletion of the D20S108 locus on 20q12 and the gain of chromosome 8 using fluorescence in situ hybridization (FISH). Genomic copy number changes are frequent in myeloid disorders such as myelodysplastic syndrome (MDS) and acute my-eloid leukemia (AML).[1-3] Deletion of the D20S108 locus is observed in 0.6 - 5% of de novo MDS patients and in less than 2% of de novo AML patients.[1,2] In MDS patients, deletion of the D20S108 locus is associated with a good prognosis[3] whereas in AML patients it is a marker of either an intermediate or an unfavorable outcome.[2] The trisomy of chromosome 8 is observed as a sole abnormality in ~5% of MDS patients or as part of a complex karyotype in >15% of such patients and is generally associated with an intermediate prog-nosis.[2,3] Additionally, trisomy 8 can be observed as a sole aberration in 5% of de novo AML patients or simultaneously with other aberrations in 15% in such patients.[2] However, trisomy 8 is more common in de novo AML cases than in therapy related AML (t-AML) with an occurrence of 7.4% vs. 3.3%, respectively[4,5] and has been associated with an intermediate prognosis.[2,3]

references1. Sole, F., et al. Haematologica, 2005. 90(9):

p.1168-78.2. Heim, S., Mitelman, F. (Ed) Cancer Cytogenetics,


3. Haase, D. Ann Hematol, 2008. 87(7):p.515-26.4. Qian, Z., et al. Chem Biol Interact, 2010. 184(1-

2): p.50-7.5. Mauritzson, N., et al. Leukemia, 2002. 16(12):

p.2366-78.

breast

lungColorectal

The EGFR/Cen7 DNA-FISH Probe is designed to detect an increase in copy number of the EGFR gene on 7p11 (previously assigned to band 7p12) rela-tive to the control Cen7 probe using fluorescence in situ hybridization (FISH) in formalin-fixed, paraffin-embedded (FPPE) tissues.[1] The EGFR gene en-codes a transmembrane protein involved in cell proliferation.[2,3] Increased copy number of the EGFR gene has been reported in non-small cell lung cancer (NSCLC) cases and some other carcinomas such as colorectal, head and neck, and breast cancer cases.[3-6] An increased EGFR copy number is predictive of response to anti-EGFR therapies.[1-5]

references1. Varella-Garcia, M. et al. J Clin Pathol, 2009. 62:970-

977.2. Toschi, L., and Cappuzzo, F. The Oncologist, 2007.

12:211-220.3. Bhargava, R. et al. Modern Patholy,2005. 18:

1027-1033.4. Hirsch, F.R. et al. J Clin Oncol, 2008. 26(20):3351-

3357.5. Personeni, M. et al. Clin Cancer Res, 2008.

14(18):5869-5876.6. Zimmermann, M., et al. Radiation Oncology, 2006.

1:11.

20

8q11

20q12

D20S108

~302 kb

telomere centromere

8

5’EGFR

3’

~ 495 kb

7p11

7

centromeretelomere

Cen7

D20

S108

/8q1

1EG

FR/C

en7


Page 17

EGR1/5p15Two Color, Enumeration ProbeRef: 11-004

ERBB2/Cen17Two Color, Enumeration ProbeRef: 22-003

AMl MDS

The EGR1/5p15 DNA-FISH Probe is designed to detect the deletion of the EGR1 gene located on 5q31 relative to the control locus 5p15 by fluores-cence in situ hybridization (FISH). The deletion of the EGR1 gene is detected in 10-15% of de novo myelodyplastic syndrome (MDS) and acute myeloid leukemia (AML) patients, and in 35-42% of therapy-related MDS (t-MDS) and therapy-related AML (t-AML) patients.[1, 2] When observed as the sole chromosomal aberration in cases of MDS (also called 5q-syndrome), dele-tion of the EGR1 gene is associated with a favorable prognosis and good response to lenalidomide treatment.[3] In cases of MDS/AML and t-MDS/t-AML, deletion of EGR1 as part of a complex karyotype is associated with a worse prognosis and unfavorable outcome.[3,4]

references1. Herry, A., et al. Eur J Haematol, 2007. 78(6): p.

457-67.2. Schoch, C., et al. Genes Chromosomes Cancer,

2002. 35(1): p. 20-9.3. List, A., et al. N Engl J Med, 2006. 355(14): p.

1456-65.4. Haase, D. Ann Hematol, 2008. 87(7): p. 515-26.

breast

The ERBB2/Cen17 DNA-FISH Probe is designed to detect the amplification of the ERBB2 gene (also named HER2/neu) on chromosome 17q12 relative to the control Cen17 using fluorescence in situ hybridization (FISH) in forma-lin-fixed, paraffin-embedded (FFPE) breast cancer tissues. Overexpression of the ERBB2 gene occurs in 25-30% of human breast carcinomas, and ~90-95% of these cases result directly from gene amplification.[1] Patients showing such an amplification are at high-risk for relapse and lower overall survival.[1-3] Amplification of the ERBB2 gene predicts a favorable response to certain chemotherapy regimens and selective monoclonal antibody ther-apy with trastuzumab.[1-5] ERBB2 amplification is also seen in other solid tu-mors such as gastric, esophageal, gynecologic, bladder, and non-small cell lung cancer and correlates with a poor prognosis.[6]

references1. Pauletti, G., et al. J Clin Oncol, 2000. 18(21):3651-64.2. Harries, M., et al. Endocr Relat Cancer, 2002.

9(2):75-85.3. Kallioniemi, O. P., et al. Proc Natl Acad Sci USA,

1992. 89(12):5321-5.4. Slamon, D. J., et al. Science, 1987. 235(4785):177-

82.5. Wolff, et al. J Clin Oncol, 2007. (1):118-145.6. Mano, M. S., et al. Cancer Treat Rev, 2007.

33(1):64-77.

5’ 3’5p15

~222 kb

5

telomere telomereEGR1

~554 kb

ERBB2

17q12 5’ 3’

~ 160 kb

centromere telomere

17Cen17

EGR

1/5p

15ER

BB

2/C

en17

Page 18

FGFR3/IGHTwo Color, Two Fusion Translocation ProbeRef: 16-005

MM

FHACttM

Four Color, Enumeration ProbeRef: 25-002The FISH-based HPV-Associated Cancer Test (FHACTTM) DNA-FISH Probe is designed to detect changes in copy number of the TERC gene located on 3q26, the D5S2095 locus located on 5p15, the D20S911 locus located on 20q13, and the Cen7 region by fluorescence in situ hybridization (FISH). Increased copy number of these genomic regions have been observed in cervical carcinoma and their precursor lesions.[1-4] This probe may be useful in identifying high-risk dysplastic lesions that progress to invasive cervical cancer.[3,4]

Cervical

The IGH/FGFR3 translocation probe is designed to detect the translocation between the FGFR3 gene located on 4p16 and the IGH gene located on 14q32 using fluorescence in situ hybridization (FISH). Rearrangement of the FGFR3 and IGH genes is designated as t(4;14) and it has been observed in ~15% of multiple myeloma (MM) patients. Detection of the t(4;14) transloca-tion is clinically relevant because it confers an aggressive phenotype with a poor prognosis and a rapid relapse after high-dose chemotherapy.[1-3]

references1. Chen, L., et al. Exp Oncol, 2007. 29(2):116-20.2. Moreau, P., et al. Blood, 2002. 100(5):1579-83.3. Avet-Loiseau, H., et al. J Clin Oncol, 2012. 30(16):

1949-52.

references1. Andersson, S., et al. Br J Cancer, 2006.

95(3):331-8.2. Rao, P.H., et al. BMC Cancer, 2004. 4:5-14.3. Narayan, G., et al. Genes Chrom Cancer, 2007.

46(4):373-84.4. Scotto, L., et al. Mol Cancer, 2008. 7:58.

FGFR33’4p16

~593 kb

telomere centromere5’

~644 kb

3’14q32

~561 kb ~420 kb

5’C segments J s

egme

nts

D se

gmen

ts

V segmentsIGH

centromere telomere

14

4

~601 kb

D5S2095centromeretelomere 5p15

~904 kb

TERCtelomere centromere3q26

~493 kb

D20S911telomere centromere20q13

3 5

207Cen7

FGFR

3/IG

HFH

AC

ttM


Page 19

IGH break ApartTwo Color, Break Apart ProbeRef: 13-014

IGH/BCL2Two Color, Two Fusion Translocation ProbeRef: 18-001The IGH/BCL2 DNA-FISH Probe is designed to detect the translocation be-tween the IGH gene located on 14q32 and the BCL2 gene located on 18q21, using fluorescence in situ hybridization (FISH). The translocation between the IGH and BCL2 gene is designated as t(14;18)(q32;q21) and is the hall-mark of follicular lymphoma (FL). The t(14;18) translocation is more frequent in lower FL grades, such as FL grades 1 and 2 (88%), than in higher grades, such as FL 3b (4-13%).[1] The translocation is also detected in 30% of diffuse large B-cell lymphoma (DLBCL) cases, but is less frequently detected in other non-Hodgkin lymphomas.[1,2] By conventional cytogenetics, the t(14;18)(q32;q21) translocation that involves the IGH and BCL2 genes is indistin-guishable from the t(14;18)(q32;q21) translocation that involves the IGH and MALT1 genes. Such translocations can be distinguished by using the re-spective DNA-FISH probes.[1]


2009 (3rd Edition). Wiley-Blackwell, New Jersey. P. 297-358.

2. Fan, Y.S., Rizkalla, K. Cancer Genet Cytogenet, 2003. 143(1): p. 73-9.

NHl

MM NHl

The IGH Break Apart DNA-FISH Probe is designed to detect the transloca-tion involving the immunoglobulin heavy chain (IGH) locus on chromosome 14q32 using fluorescence in situ hybridization (FISH). At least 40 transloca-tion gene partners to the IGH locus have been identified.[1] Rearrangements involving the IGH locus and specific partners are mainly found in multiple myeloma (MM)[2,3] and non-Hodgkin’s lymphoma (NHL) subtypes.[1,4] The prognosis is dependent upon the translocation partner and the type of malig-nancy. The design of the IGH Break Apart DNA-FISH Probe allows the visu-alization of a break between the constant C domain (red) and the variable V domain (green) of the IGH locus and the resulting translocation.

references1. Bernicot, L., et al. Cytogenet Genome Res, 2007.

118(2-4):345-52.2. Moreau, P., et al. Blood, 2002. 100(5):1579-83.3. Avet-Loiseau, H., et al. Blood, 2002. 99(6):2185-91.4. Haferlach, C., et al. Leukemia, 2007. 21(12):2442-51.

All

All

3’ 5’18q21

BCL2

~441 kb

telomere centromere

~575 kb

18

14

3’14q32

~561 kb ~420 kb

5’C segments J s

egme

nts

D se

gmen

ts

V segmentsIGH

centromere telomere

3’14q32

~561 kb ~420 kb

5’C segments J s

egme

nts

D se

gmen

ts

V segmentsIGH

centromere telomere

14

IGH

/BC

L2IG

H b

reak

Apa

rt

Page 20

IGH/MAFTwo Color, Two Fusion Translocation ProbeRef: 16-010

IGH/MALT1Two Color, Two Fusion Translocation ProbeRef: 17-004

MM

NHl

The IGH/MAF DNA-FISH Probe is designed to detect the translocation be-tween the IGH gene located on 14q32 and the MAF gene located on 16q23 by fluorescence in situ hybridization (FISH).[1] The translocation between the IGH and MAF gene, designated as t(14;16)(q32;q23), is found in 2-10% of multiple myeloma (MM) cases and is associated with a more aggressive dis-ease along with an unfavorable prognosis and outcome.[2,3]

references1. Chesi, M., et al., Blood, 1998. 91(2): p. 4457-63.2. Kapoor, P., et al. Mayo Clin Proc, 2010. 85(6): p.

532-7.3. Boyd, K.D., et al., Leukemia, 2012. 26: p. 349-55.

The IGH/MALT1 DNA-FISH probe is designed to detect the translocation between the IGH gene located at 14q32 and the MALT1 gene located at 18q21 by fluorescence in situ hybridization (FISH), designated as t(14;18)(q32;q21). The rearrangement of IGH/MALT1 has been observed in 10-20% of mucosa-associated lymphoid tissue (MALT) lymphoma, predominantly occurring in liver, skin, and ocular adnexa.[1-3] By conventional cytogenetics, t(14;18)(q32;q21) involving the IGH and MALT1 genes is indistinguishable from the t(14;18)(q32;q21) involving the IGH and BCL2 genes (the hallmark of follicular lymphoma). These translocations can be distinguished by FISH, using the respective DNA-FISH probes.

references1. Streubel, B., et al. Blood, 2003. 101(6): p.2335-9.2. Streubel, B., et al. Leukemia, 2004. 18(10):

p.1722-6.3. Murga Penas, E.M, et al., Leukemia, 2003. 17(11):

p. 2225-9

16

3’ 5’16q23

MAF

~655 kb

telomere centromere

14

3’14q32

~561 kb ~420 kb

5’C segments J s

egme

nts

D se

gmen

ts

V segmentsIGH

centromere telomere

18

5’ 3’18q21

MALT1

~738 kb

telomere centromere

~641 kb

14

3’14q32

C segmentsV segments

centromere telomere

~561 kb ~420 kb

5’

IGH

J seg

ment

s D

segm

ents

IGH

/MA

FIG

H/M

ALT

1


Page 21

MDM2/D12S1837Two Color, Enumeration ProbeRef: 14-017

MLL break ApartTwo Color, Break Apart ProbeRef: 11-002

Cll NHl

The MDM2/D12S1837 DNA-FISH Probe is designed to detect both the poly-ploidy of chromosome 12 and the amplification of the MDM2 gene located on 12q14.3-q15 relative to the control locus D12S1837 located on 12p11 by fluo-rescence in situ hybridization (FISH). MDM2 is frequently amplified in many solid tumor, including soft tissue tumors. Well-differentiated liposarcomas has been found by molecular and cytogenetic studies to contain amplification of the 12q13-15 region, including the MDM2 gene.[1,2] Precise recognition of be-nign lipoma and well-differentiated liposarcoma by core needle biopsy can facilitate appropriate clinical management.[3] Trisomy of chromosome 12 (+12) is a commonly observed numerical aberration in non-Hodgkin lymphomas (NHL) cases. It has been observed in ~15-50% chronic lymphocytic leukemia (CLL) cases.[4] In CLL, trisomy 12, when observed as a sole abnormality, ap-pears to have limited prognostic implications and is often associated with an atypical morphology, although it is only observed in a subset of tumor cells.[5] references

1. Weaver, J., et al. Modern Pathol, 2008. 21(8): p. 943-9.

2. Dal Cin, P., et al. Cancer Genetics and Cytogenetics, 1993. 68(2): p. 85-90.

3. Weaver, J., et al., Modern Pathol, 2010. 23(10): p. 85-90.

4. Heim, S., Mitelman F. (Ed) Cancer Cytogenetics, 2009 (3rd Edition). Wiley-Blackwell, New Jersey. P. 310.

5. Dohner, H., et al. J Mol Med, 1999. 77(2):p.266-81.

All AMl

The MLL Break Apart probe is designed to detect the translocation involving the MLL gene on chromosome 11q23 using fluorescence in situ hybridiza-tion (FISH). At least 104 translocation partner genes have been identified.[1] Translocation of MLL is found in ~3-10% of acute lymphoblastic leukemia (ALL) cases, and in ~8-10% of acute myeloid leukemia (AML) cases, and is prognostically relevant in these leukemias.[2,3] However, the prognostic impli-cation is dependent on the age and phenotype of the leukemia. MLL rear-rangement has been observed in ~80% of infant ALL cases and is associat-ed with a high risk in such cases and requires aggressive treatment. In AML, the prognosis is intermediate regardless of age. MLL translocations are also found in ~25% of patients with therapy-related leukemias, particularly fol-lowing treatment with DNA topoisomerase II inhibitors and the prognosis in such patients is poor.[2,3] In addition to translocations, deletions of 3’ MLL and amplification of MLL also occurs in a subset of ALL and AML cases.[4,5] references

1. Meyer, C., et al., Leukemia, 2009. 23: 1490-9.2. Coenen, E.A., et al., Blood, 2011. 117(26): 7102-11.3. Chowdhury, T., et al. Blood Cells Mol Dis, 2008.

40:192-199.4. Barber, K. E, et al. Genes Chromosomes Cancer,

2001. 41:226-271.5. Andersen, M. K, et al. Genes Chromosomes Cancer,

2001. 31:33-41.

12

D12S1837

~304 kb

telomere telomere 5’ 3’MDM2

~289 kb

telomerecentromereMLL

5’ 3’11q23

~770 kb ~820 kb

11

MD

M2/

D12

S183

7M

LL b

reak

Apa

rt

Page 22

MYB/SHgC-79576Two Color, Enumeration ProbeRef: 14-016

MYC break ApartTwo Color, Break Apart ProbeRef: 13-008

The MYB/SHGC-79576 DNA-FISH Probe is designed to detect copy number changes of the MYB locus located on 6q23 relative to a 6p12 control locus, using fluorescence in situ hybridization (FISH).[1] MYB gene is involved in duplications, translocation, and deletions in a variety of cancer types. Loss of MYB has been observed in ~5% of chronic lymphocytic leukemia (CLL) cases[2] and has been associated with a poor prognosis in CLL cases.[2] In breast tumors, the gain of the MYB gene has been observed in ~ 30% of hereditary BRCA1 positive breast tumors.[3]

references1. Barletta, C., et al., Science, 1987. 235(4792): p.

1064-7.2. Reddy, K. S. Br J Haematol, 2006. 132(6):

p.705-22. 3. Kauraniemi, P., et al. Cancer Res, 2000. 60(19):

p.5323-8.

The MYC Break Apart DNA-FISH Probe is designed to detect the translo-cation between the MYC gene located at 8q24 and one of 11 known trans-location partner loci using fluorescence in situ hybridization (FISH). The most common translocation, t(8;14)(q24;q32), is found in 75-85% of Burkitt lymphoma (BL) cases and is the cytogenetic hallmark of BL.[1,2] The t(8;14)(q24;q32) in BL is associated with an aggressive clinical course that responds well to high-intensity, brief-duration drug regimens with an overall favorable outcome.[2,3] Translocation of MYC is often detected as a secondary genomic abnormality at low frequencies in high-grade B-cell lymphomas, such as dif-fuse large B-cell lymphoma (DLBCL) (5-16%) and chronic lymphocytic leu-kemia (CLL) (0.1-2%).[1,3-5] In DLBCL, the presence of the MYC translocation is associated with an aggressive disease with a poor prognosis and an un-favorable outcome.[4] MYC translocation has also been observed in 4-6% of acute lymphoblastic leukemia (ALL).[6] references

1. Heim, S., Mitelman, F. (Ed) Cancer Cytogenetics, 2009 (3rd Edition). Wilwy-Blackwell, New Jersey. p. 326-328.

2. Blum, K. A., et al. Blood, 2004. 104(10): p. 3009-20.3. Lones, M. A., et al. J Pediatr Hematol Oncol, 2004.

26(3): p. 169-78.4. Snuderl, M., et al. Am J Surg Pathol, 2010. 34(3):

p. 327-40.5. Aukema, S.M., et al., Blood, 2011. 17(8): p. 2319-31.6. Moorman, A. V., et al. Blood, 2010. 115(2): p. 206-14.

All

NHl

All Cll

Cll

5’ 3’SHGC-79576

~777kb

6

telomere telomereMYB

~494kb

8

5’ 3’8q24

MYC

~647 kb

telomere centromere

~594 kb

MYB

/SH

gC

-795

76M

YC b

reak

Apa

rt


Page 23

MYC/IGHTwo Color, Two Fusion Translocation ProbeRef: 13-004

MYH11/CBFBTwo Color, Two Fusion Translocation ProbeRef: 12-010

All NHl

The MYC/IGH DNA-FISH Probe is designed to detect the translocation be-tween the MYC gene located at 8q24 and the IGH gene located at 14q32, us-ing fluorescence in situ hybridization (FISH). The translocation between the MYC and IGH gene is designated as t(8;14)(q24;q32) and is the cytogenetic hallmark of Burkitt lymphoma (BL), which is found in 75-85% of patients.[1]

In BL, t(8;14)(q24;q32) is associated with an aggressive clinical course that responds well to high-intensity, brief-duration drug regimens with an overall favorable outcome.[1,2] This rearrangement has been observed at lower fre-quencies in other non-Hodgkin lymphomas (NHLs), such as diffuse large B-cell lymphoma (DLBCL) (<10%).[3,4] This translocation also occurs less fre-quently in acute lymphoblastic leukemia (ALL) where it has been associated with an unfavorable outcome.[3,5]

references1. Blum, K. A., et al. Blood, 2004. 104(10): p.3009-20.2. Lones, M. A., et al. J Pediatr Hematol Oncol, 2004.

26(3): p.169-78.3. Heim, S., et al. Cancer Cytogenetics, 2009 (3rd

Edition).4. Akasaka, T., et al. J Clin Oncol, 2000. 18(3):

p.510-18.5. Moorman, A. V., et al. Blood, 2010. 115(2):

p.206-14.

AMl

The MYH11/CBFB DNA-FISH Probe is designed to detect the pericentric inversion of chromosome 16 inv(16)(p13q22) and t(16;16)(p13;q22) involving the MYH11 gene on 16p13 and the CBFB gene on 16q22 using fluorescence in situ hybridization (FISH). The rearrangement of the MYH11 and CBFB gene results in a fusion of both genes. The inv(16) abnormality is found in ~5 - 8% of all of de novo acute myeloid leukemia (AML) cases and is associ-ated with AML-M4eo subtype (based on FAB classification).[1,2] Inv(16) and t(16;16) has also been observed in therapy related myelodysplastic syndrome (t-MDS) cases and in eosinophilic blast crisis of chronic myelogenous leuke-mia (CML) cases.[2] Whether alone or as part of a complex patient karyotype, inv(16) is indicative of a good prognosis in AML cases.[3]

references1. LeBeau, M.M., et al., N Eng J Med, 1983. 309(11):

630-6.2. Huret, J. L. inv(16)(p13q22),t(16;16)(p13;q22),

del(16)(q22). www.AtlasGeneticsOncology.org.3. Mrozek K, et al. Blood Rev, 2004. 18:115-136.

MDSCMl

8

5’ 3’8q24

MYC

~647 kb

telomere centromere

~594 kb

14

3’14q32

~561 kb ~420 kb

5’C segments J s

egme

nts

D se

gmen

ts

V segments

IGH

centromere telomere

MYH113’ 5’16p13

~472 kb ~233 kb

CBFB5’ 3’16q22

~444 kb ~472 kb

centromere telomere

telomere centromere

16

MYC

/IGH

MYH

11/C

BFB

Page 24

PBX1/E2ATwo Color, Two Fusion Translocation ProbeRef: 13-001

PML/RARATwo Color, Two Fusion Translocation ProbeRef: 12-008

All

The PBX1/E2A (also named TCF3/PBX1) DNA-FISH Probe is designed to detect the translocation between the PBX1 gene located on 1q23 and the E2A gene located on 19p13, using fluorescence in situ hybridization (FISH). The translocation between the PBX1 and E2A gene is designated as t(1;19)(q23;p13) and occurs in ~ 6% of pediatric and adult acute lymphoblastic leu-kemia (ALL) cases; as determined by conventional cytogenetics and reverse transcription-polymerase chain reaction.[1] In both pediatric and adult ALL, the translocation is correlated with a negative prognosis.[2,3] It may occur as a balanced translocation, t(1;19)(q23;p13), or as an unbalanced translo-cation, der(19)t(1;19)(q23;p13), where only the derivative chromosome 19 is present.[2] The unbalanced translocation, der(19), is the most common form and accounts for 75% of all PBX1/E2A rearrangements.[2] Both balanced and unbalanced translocations are sometimes observed in the same patient as separate clones.[2]

references1. Hunger, S.P., et al., Blood, 1991. 77(4): p. 687-93.2. Shearer, B. M., et al. Br J Haematol, 2005. 129(1):

p.45-52.3. Anderson, M.K., et al., Br J Haematol, 2011:

155(2): p. 235-43.

AMl

The PML/RARA DNA-FISH Probe is designed to detect the translocation between the PML gene on chromosome 15q24 (previously assigned to band 15q22) and the RARA gene on chromosome 17q21, using fluorescence in situ hybridization (FISH). The t(15;17) translocation is the diagnostic hallmark of acute promyelocytic leukemia (APL), a sub-group of acute myelogenous leukemia (AML), and results in the fusion of the PML and RARA genes.[1] The presence of a PML-RARA fusion predicts a favorable response to differen-tiation therapy with all-trans retinoic acid (ATRA) and is currently the most curable subtype of acute myeloid leukemia (AML).[1-3] The t(15;17) transloca-tion has also been identified in chronic myeloid leukemia (CML) cases with promyelocytic blast crisis.

references1. Kakizuka, a., et al., Cell, 1991. 66: 663-74.2. Brockman, S. R, et al. Cancer Genet Cytogenet,

2003. 145:144-15.3. Mistry, A.R., et al., Blood Rev., 2003. 17(2): 71-97.

CMl

19

1q23

PBX1

~326 kb

telomere centromere

3’ 5’19p13

E2A

~307 kb

centromere telomere

~255 kb

~275kb

1

5’ 3’

15q24

telomerecentromere17q21

~570 kb ~510 kb

centromere telomere

~270 kb ~350 kb

PML

5’ 3’

RARA

5’ 3’

15

17

PBX1

/E2A

PML/

RA

RA


Page 25

AMl CllRB1/D13S1009Two Color, Enumeration ProbeRef: 27-014

TP53/RARATwo Color, Enumeration ProbeRef: 14-015The TP53/RARA DNA-FISH Probe is designed to detect the deletion of the TP53 gene located on 17p13, relative to the control marker gene RARA lo-cated on 17q21, using fluorescence in situ hybridization (FISH). The TP53 gene is a known tumor suppressor gene and is frequently deleted in a wide variety of solid tumors [1, 2] and hematologic malignancies such as mature B-cell neoplasms,[3] myeloid disorders such as acute myeloid leukemia (AML),[4] and myelodysplastic syndrome (MDS).[5] Deletion of the TP53 gene has been associated with advanced stage, shortened survival, and resistance to treat-ment in several malignancies and solid tumors.[3,6] The loss of TP53 is also found in chronic lymphocytic leukemia (CLL) cases and is associated with a very poor clinical outcome.[7,8]

references1. Bertheau, P., et al. Pathobiology, 2008 (Review).

75(2):132-9.2. Doak, S. H, et al. Br J Cancer, 2003. 89(9): 1729-35.3. Döhner, H., et al. J Mol Med, 1999 (Review). 77(2):

266-81.4. Seifert, H., et al. Leukemia, 2009. 23(4): 656-63.5. Silveira, C. G., et al. Leuk Res, 2009. 33(1): 19-27.6. Hof, J., et al. J Clin Oncol, 2011. 29(23): 3185-93.7. Dal Bo, M., et al. Genes Chromosomes Cancer,

2011. 50(8): 633-43.8. Boyd, K.D., et al., Leukemia, 2012. 26: 349-55.

The RB1/D13S1009 DNA-FISH Probe is designed to detect loss of the RB1 gene on chromosome 13q14 relative to the control marker, D13S1009 on chromosome 13q34, using fluorescence in situ hybridization (FISH). The RB1 gene is a well characterized tumor-suppressor gene and bi-allelic in-activation of the gene due to mutations and/or deletions is causal for the development of Retinoblastoma (RB). Deletion of the RB1 gene is also com-mon in a wide variety of solid tumors and hematologic malignancies such as chronic lymphocytic leukemia (CLL), multiple myeloma (MM), acute myelo-cytic leukemia (AML), myelodysplastic syndrome (MDS), and chronic myelo-proliferative disorders.[1-3] The RB1 gene is proximal to the D13S25 locus at 13q14, which is often co-deleted with the RB1 gene in some B-cell hemato-logic malignancies.

references1. Lee, W.-H., et al., Science, 1987. 235: 1394-99.2. Steigenbauer, S., et al., Blood, 1993. 81(8):

2118-24.3. Dao, D.D., et al., Leukemia, 1994. 8(8): 1280-4.

MM

MDSCll

breast

lung

Colorectal

MDS

AMl

centromere telomere

13

~305 kb ~368 kb

RB15’ 3’

D13S1009

RARA5’

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TP535’ 3’

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sCorinG GuidanCe & siGnal interPretationSignal interpretation depends on the DNA-FISH Probe type and should be made with the full knowl-edge of the product design. The following table divides the DNA-FISH Probe into two types: transloca-tion and enumeration probes.

The true green signal and the green artifact are indistinguish-able. The green artifact or fluorescent debris is so bright, it interferes with a proper evalu-ation.

The signals are indistinguishable from the background.

The irregular red signal and the green signal on the periphery could be artifacts.

Nucleus is physically damaged and nuclei overlap prevents distinction of which signals be-long to which nucleus.

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transloCation Probes

Count as 1R2G1F signals.

Count as 2F signals. Depend-ing on the overlap, the red and green signal overlap can ap-pear as yellow.

Count as 2F signals; the dis-tance between the red and green signals is less than one signal width apart.

Count as 1R1G2F signals, depending on the probe this pattern could be indicative of a reciprocal translocation.

The boundary of each nucleus is clear; for each nucleus count as 2R2G signals.

Count as 2R2G signals; the green signal is a split.

enuMeration Probes

The boundary of each nucleus is clear; for each nucleus count as 2R2G signals.

Count as 2R2G signals; the green signal is a split.

Count as 2R2G signals; the red and green signals are less than one signal width apart. When overlapping occurs, the signal appears as yellow.

Count as >6R2G signals; one red signal is focally amplified.

Count as 1R2G signals; the pattern is indicative of the de-letion of the red locus.

**Characteristic of certain probes that target a highly transcribed region (i.e. IGH).

Count as 2R2G signals; the green signal is stringy or dis-persed.**

Count as 2R2G signals, de-pending on the nuclear orga-nization, the red and green signal can overlap and appear as yellow.

Count as 1R1G signals; the pat-tern is indicative of the loss of an entire chromosome.

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filter sPeCifiCations

The following table shows the filter requirements for Cancer Genetics Italia DNA-FISH Probes.

1. Applies to all the CGI Italia DNA-FISH Probes.

2. Preferably used for three or more color DNA- FISH Probes.

filter MaintenanCe reCoMMendationFilters age with use and should be regularly examined for dirt and/or imperfections which may occur over time. Heat from the high intensity transmitted light can etch and damage frequently used filters (such as the DAPI filter), resulting in dimmer images and unevenly illuminated areas. Such filters should be replaced. Handle filters according to manufacturers recommendations.

troublesHootinG1. I am observing weak signals. How can I correct this?

Weak signals may be observed due to:• Use of an old mercury lamp (run time >200 hours); replace the mercury lamp after 200 hours of run time. • Use of unsuitable immersion oil; use immersion oil with no auto-fluorescence in the UV range. • Use of unsuitable filters; use the appropriate filter sets. Also, check that the excitation and emission of

the dyes fall within the range claimed by the filter manufacturer. The filter specifications table (see table above) identifies the filter requirements for Cancer Genetics Italia DNA-FISH Probes. Also, be sure the filters are well maintained and have no visible damage.

• Inadequate denaturation of the DNA-FISH Probe; increase the denaturation temperature and/or duration. Optionally, the specimen may be treated with pepsin prior to denaturation.

• Insufficient pretreatment of the specimen (visible cytoplasm on the specimen appearing as a green or yellow haze). In such cases it may be beneficial to perform the optional pretreatment steps outlined in the Instructions for Use (IFU).

• Baking or aging the slide for too long; check the temperature of the hot plate or oven that is used to age the slide. The time and temperature routinely used to artificially age the slide varies among specimen types and laboratories. Adjust conditions according to laboratory SOPs.

2. Why do I see areas without signals?Areas without signals indicate a lack of hybridization in those areas. This may be due to:

• Presence of air bubbles during hybridization, or application of an insufficient volume of Probe.• Before adding the coverslip, gently remove all air bubbles with a pipette tip or needle. After adding the

coverslip, gently remove any air bubbles by rolling a pencil eraser evenly across the coverslip. Also, ensure there is sufficient Probe volume (10 μl Probe/22×22mm target area). Increase or decrease the volume of Probe and size of coverslip proportionally with any increase or decrease in the size of the target area.

• Low permeability of the specimen to the Probe, which results in resistance to hybridization. Optionally, the specimen may be treated with pepsin prior to denaturation (protocol is provided in the IFU).

Fluorophore excitationmax emissionmax

green 496 nm 520 nm

red1 580 nm 603 nm

red2 593 nm 612 nm

DAPI 360 nm 460 nm

gold 525 nm 551 nm

Aqua 431 nm 480 nm

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3. Why do the signals appear to be fading?The signals may be fading due to:

• Photobleaching of the slide; always close the microscope shutter when not observing the specimen. It is important that hybridized slides are stored at 4°C and protected from light. Fluorescently labeled probes are readily photobleached by exposure to light, both before and after hybridization. To minimize photo-bleaching, handle Probes and hybridized specimens in reduced light as much as possible.

• Use of immersion oil with auto-fluorescence in the UV range. Be sure to use immersion oil with no auto-fluorescence in the UV range.

• Use of expired or oxidized Antifade/DAPI, which will appear deep purple/brown in color. Verify the expira-tion date of the Antifade used.

• Image acquisition. Some photobleaching will occur from extended exposure to the UV source during im-age acquisition.

4. Why do I see no signals on my hybridized slide?Seeing no signals on a hybridized slide indicates a lack of hybridization. This may be due to:

• Application of an insufficient Probe volume and/or lack of cells in target area. It is good practice to check the slide under a phase contrast microscope, prior to hybridization, to select appropriate areas for hy-bridization.

• Inappropriate denaturing/incubation conditions. Check the temperature of the controlled hot plate prior to use and ensure that the hot plate is calibrated regularly. Make sure to respect the recommended time and temperature for denaturation/incubation, and ensure that the incubation chamber is humidified, as recommended in the IFU (provided with the product and available at www.cancergeneticsitalia.com).

• Too much cytoplasm present (appears as a green or yellow haze), which impedes the Probe from hy-bridizing to the target DNA. Optionally, the specimen may be treated with pepsin prior to denaturation (protocol is provided in the IFU).

• Specimen quality not optimal for FISH. Occasionally a specimen will not lend itself to be processed for FISH. This may be the case in a subset of formalin-fixed paraffin embedded (FFPE) specimens when a variety of involved tissues (such as bone) may inhibit processing of the slide, and yield poor FISH signals, or no signals at all.

5. Why is the nuclear morphology compromised?The nuclear morphology may be compromised due to:

• Over-denaturation of the specimen; reduce the duration and/or temperature of denaturation. It is impor-tant to check the temperature of the hot plate or oven used to age the slide.

• Excessive pretreatment of the specimen, which leads to chromatin loss; reduce the pepsin concentration and/or incubation time during pretreatment, or eliminate this optional step altogether.

6. Why do I see a high level of background noise?A high level of background noise may be due to:

• Use of an unclean slide during specimen preparation; clean slide with 70% ethanol and wipe with a lint free cloth prior to dropping the specimen on the slide.

• Presence of cellular debris in the specimen; wash the specimen pellet in fresh fixative prior to dropping. Allow residual debris to settle to bottom of tube before drawing up the specimen.

• Allowing the slide to dry during hybridization or post-hybridization washes; when placing coverslip over target area, ensure proper sealing with rubber cement to prevent the Probe from drying. Also, do not al-low slide to dry after removing coverslip for post-hybridization washes, or in between the washes.

• Inadequate post-hybridization washing; ensure the wash buffers have reached the optimal temperature recommended in the IFU prior to starting the washes. Also be sure to wash the slide for the duration recommended in the IFU.

7. Why do I see a high level of nuclear background noise?A high level of nuclear background noise may be caused by:

• Inadequate post-hybridization washing; although rarely observed, a high level of nuclear background noise may be reduced by increasing the washing stringency and/or duration.

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Cancer genetics Italia2013 DNA-FISH Probe Catalogwww.cancergeneticsitalia.com

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Cancer genetics Italia2013 DNA-FISH Probe Catalogwww.cancergeneticsitalia.com

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orderinG inforMation

CGI Italia’s DNA-FISH Probes are available in a ready to use, 10 tests per vial format.

To place an order, please contact your local authorized distributor. To locate your local authorized distribu-tor please visit our website at www.cancergeneticsitalia.com; if a distributor is not located within your region, please contact us at [email protected].

We are dedicated to providing professionals the most clinically relevant products and are constantly devel-oping new reagents. If you would like more information about our pipeline or if you have specific inquiries, please contact us at [email protected].

If you are interested in distributing our DNA-FISH Probes or would like more information, please contact us at [email protected].

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ProduCt index by disease

disease Probe naMe referenCe nuMber PaGe

HeMAtologIC ProbeSAlCl ALK Break Apart 21-002 11

ALK/NPM1 21-001 12

All A20/PRDM1/SHGC-79576 19-001 10ABL1/BCR 10-001 11IGH/BCL2 18-001 19IGH Break Apart 13-014 19MLL Break Apart 11-002 21MYB/SHGC-79576 14-016 22MYC Break Apart 13-008 22MYC/IGH 13-004 23PBX1/E2A 13-001 24

AMl AML1/ETO 12-005 12D7S486/Cen7 11-007 15D20S108/8q11 11-001 16EGR1/5p15 11-004 17MLL Break Apart 11-002 21MYH11/CBFB 12-010 23PML/RARA 12-008 24RB1/D13S1009 27-014 25TP53/RARA 14-015 25

Cll ATM/D11S1251 14-018 13D13S25/D13S1009 14-009 15MDM2/D12S1837 14-017 21MYB/SHGC-79576 14-016 22MYC Break Apart 13-008 22RB1/D13S1009 27-014 25TP53/RARA 14-015 25

CMl ABL1/BCR 10-001 11MYH11/CBFB 12-010 23PML/RARA 12-008 24

MM 5p15, 9q34, 15q24 16-003 10CCND1/IGH 14-006 14

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ProduCt index by disease

disease Probe naMe referenCe nuMber PaGe

MM D13S25/D13S1009 14-009 15FGFR3/IGH 16-005 18IGH Break Apart 13-014 19IGH/MAF 16-010 20RB1/D13S1009 27-014 25

MDS D7S486/Cen7 11-007 15D20S108/8q11 11-001 16EGR1/5p15 11-004 17MYH11/CBFB 12-010 23RB1/D13S1009 27-014 25TP53/RARA 14-015 25

NHl A20/PRDM1/SHGC-79576 19-001 10API2/MALT1 17-001 13ATM/D11S1251 14-018 13BCL6 Break Apart 18-010 14CCND1/IGH 14-006 14D13S25/D13S1009 14-009 15IGH/BCL2 18-001 19IGH Break Apart 13-014 19IGH/MALT1 17-004 20MDM2/D12S1837 14-017 21MYC Break Apart 13-008 22MYC/IGH 13-004 23

SolID tuMor ProbeSbreast EGFR/Cen7 22-007 16

ERBB2/Cen17 22-003 17TP53/RARA 14-015 25

Colorectal EGFR/Cen7 22-007 16TP53/RARA 14-015 25

Cervical FHACTTM 25-002 18lung ALK Break Apart 21-002 11

EGFR/Cen7 22-007 16TP53/RARA 14-015 25

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ProduCt index by naMe

vProbe naMe referenCe # aberration PaGe

5p15, 9q34, 15q24 16-003 Hyperdiploidy 5, 9, 1510

A20/PRDM1/SHgC-79576 19-001 del(6q23),del(6q21)/6p12ABL1/BCR 10-001 t(9;22)

11ALK break Apart 21-002 t(2p23)ALK/NPM1 21-001 t(2;5)

12AML1/ETO 12-005 t(8;21)API2/MALT1 17-001 t(11;18)

13ATM/D11S1251 14-018 del(11q22)/11p15BCL6 break Apart 18-010 t(3q27)

14CCND1/IGH 14-006 t(11;14)D7S486/Cen7 11-007 del(7q31)/Cen7

15D13S25/D13S1009 14-009 del(13q14)/13q34D20S108/8q11 11-001 del(20q12) & trisomy 8

16EGFR/Cen7 22-007 EGFR AmplificationEGR1/5p15 11-004 del(5q31)/5p15

17ERBB2/Cen17 22-003 ERBB2 AmplificationFGFR3/IGH 16-005 t(4;14)

18FHACttM 25-002 3q26/5p15/20q13/Cen7IGH/BCL2 18-001 t(14;18)

19IGH break Apart 13-014 t(14q32)IGH/MAF 16-010 t(14;16)

20IGH/MALT1 17-004 t(14;18)MDM2/D12S1837 14-017 12q15/12p11

21MLL break Apart 11-002 t(11q23)MYB/SHgC-79576 14-016 6q23/6p12

22MYC break Apart 13-008 t(8q24)MYC/IGH 13-004 t(8;14)

23MYH11/CBFB 12-010 inv(16)PBX1/E2A 13-001 t(1;19)

24PML/RARA 12-008 t(15;17)RB1/D13S1009 27-014 del(13q14)/13q34

25TP53/RARA 14-015 del(17p13)/17q21

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ProduCt index by CHroMosoMe nuMber

CHroMosoMe Probe naMe referenCe nuMber PaGe

1;19 PBX1/E2A 13-001 242 ALK Break Apart 21-002 112;5 ALK/NPM1 21-001 123 BCL6 Break Apart 18-010 143, 5, 7, 20 FHACTTM 25-002 184;14 FGFR3/IGH 16-005 185 EGR1/5p15 11-004 175, 9, 15 5p15, 9q34, 15q24 16-003 106 A20/PRDM1/SHGC-79576 19-001 106 MYB/SHGC-79576 14-016 227 D7S486/Cen7 11-007 157 EGFR/Cen7 22-007 168 MYC Break Apart 13-008 228;14 MYC/IGH 13-004 238, 20 D20S108/8q11 11-001 168;21 AML1/ETO 12-005 129;22 ABL1/BCR 10-001 1111 ATM/D11S1251 14-018 1311 MLL Break Apart 11-002 2111;14 CCND1/IGH 14-006 1411;18 API2/MALT1 17-001 1312 MDM2/D12S1837 14-017 2113 D13S25/D13S1009 14-009 1513 RB1/D13S1009 27-014 2514 IGH Break Apart 13-014 1914;16 IGH/MAF 16-010 2014;18 IGH/BCL2 18-001 1914;18 IGH/MALT1 17-004 2015;17 PML/RARA 12-008 2416 MYH11/CBFB 12-010 2317 ERBB2/Cen17 22-003 1717 TP53/RARA 14-015 25

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notes

notes

Your local distributor:

© 2013 Cancer genetics Italia S.r.l.

Via San Vito n. 6 20123 Milano – Italia Web: www.cancergeneticsitalia.comemail: [email protected]: +39 02 363 11777


Documents

CGI Italia: DNA-FISH Probe Catalog